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Nanobacteria-Are they or Aren't they Alive?

BIO 181 FInal Lab Presentation

Farzana Abdulla Abdulla

on 7 December 2012

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Transcript of Nanobacteria-Are they or Aren't they Alive?

photo (cc) Malte Sörensen @ flickr Evidence of Nanobacteria Are they or aren't
they alive? NANOBACTERIA What does it mean to be alive? How would you test for this property? Experiment 1
Transferability Experiment 2
Gamma Radiation Experiment 3
Kidney Stones What are nanobacteria? Bacteria are classified as prokaryotic cells.
- Prokaryotic cells lack membrane-bound organelles, have no nucleus, and have circular DNA.
Nanobacteria are very small (about 0.2 to 0.5 micrometers).
These bacteria stimulate a process called biomineralization.
- Inorganic crystalline structures that form in association with biological macromolecules (also called calcification)
Sometimes, biomineralization occurs in the wrong areas of the body
- Ex: Kidney stones What is the Evidence that Nanobacteria Are Alive? Corroborative Evidence Data Sheet 1 For research to have substance, it needs to be reproducible. Cisar's lab group wanted to further investigate Kajandar and found evidence against nanobacteria being alive.
Culture of

Gamma Radiation


Cell-Like Appearance

DNA Staining

DNA Isolation

Protein Isolation

PCR for 16s rDNA Generated a biofilm on surface of culture plate in 3 weeks

Prevented formation of biofilm

A 1:10 dilution of the nanobacteria culture was made in a new plate and a new biofilm was formed; several months

Coccoid (round) appearance

Hoechst staining is not focused, does not appear to specifically localize on cells

No evidence of DNA based on a 260nm wavelength

Protein gel electrophoresis only show a couple of proteins

-PCR amplified DNA that was only 85% similar to the published sequence
-The same products were found in other samples w/o nanobacteria
-PCR product was 99% identical to a Pseudomonas
-Published sequence of 16s rDNA was 99% identical to 16s rDNA from Phyllobacterim Is this property present in all living things? Properties of Life Consist of one or more cells Contain and use genetic information Respond to stimuli
Adapt Regulate internal environment Yes The
Debate A Case Study of What It Means to Be a Biological Organism Conclusion Nanobacteria are not alive. - Cells must be at least 200 nm to conduct functions like DNA replication. Nanobacteria are much smaller.
- Nanobacteria do not contain DNA or RNA.
- Instead, nanobacteria contain hydroxyapatite (HPA) instead of DNA or RNA. HPA makes up teeth enamel and bones in humans and animals.
- Antibodies react to nanobacteria.
- Since there is no evidence of DNA, there are also hardly any proteins, which could have been made from the contaminant bacteria that have invaded the experimental cell culture plates.
-How can nanobacteria can grow and reproduce if there is no passing of genetic material? Viruses need genetic material to reproduce. Perhaps this may suggest that their growth process is not biological at all, but chemical, like crystal growth, which can grow under proper circumstances -Nanobacteria: discovered by Drs. E. Olavi
Kajander & Neva Ciftcioglu
-Nanobacteria are said to be: poorly
disruptable, stainable,
[These techniques didn't apply in this case]
fixable, and resistant to heat( Drs.)
-Coccoid particles from human and cow blood
were isolated and observed. Background info: Data Sheet 2 *Data sheet one exhibits light and microscopic images on nanobactera

*These images showed that nanobacteria range in size(image C)

*The nanobacteria seem to be surrounded by a hairy looking boundry (image E) *Here, nanobacteria were under SF conditions

*Images E and F show the bacteria deatching from their
culture vessel.

CITE: http://phys.org/news128167633.html#jCp What is so controversial?

How do they replicate?
Are they alive? Observe cell behavior in response to factors Isolate cells and view growth after elapsed time Place cells in different environments and see reactions Check for cells and types of organelles under microscopes *nanobacteria are
so small, that is
why their existence
is so controversial
*the pictures show that these
nanobacteria seem to undergo "cell division" "they have the ability to nucleate hydroxyapatite (HAP), a calcium phosphate crystal that largely composes the bones and teeth of humans and animals. Previous research has suggested that this might be how the nanobacteria self-replicate" Final Chapter Experiment Results Cisar Lab Conclusion Alternate Conclusion Energy use by
nanobacteria Cultures of nanobacteria were exposed to 0.1%
sodium azide - a powerful inhibitor of cellular
respiration. The formation of a biofilm continued
even in the presence of this poison. Nanobacteria would still generate a biofilm because it has a strong defense against sodium azide Since sodium azide is a powerful inhibitor of cellular
respiration and the formation of a biofilm is still present,
this shows that the nanobacteria have a high chance
of not being a living cell. This is because the
sodium azide would have inhibited the forming
of a biofilm if the cell was living. "Growth" of dilute cultures Cultures of nanobacteria were diluted to a higher
degree than that used by Kajander. Dilutions of
1:100 or 1:1000 were cultured as before. At
these high dilutions there was no evidence of
biofilm formation even after 8 weeks. The inability to form a biofilm after a higher degree of dilution than the ratio as cultured before demonstrated that the nanobacteria have a limited capacity in transferability. The nanobacteria need high concentrated transfers in order to be subcultivated and reproduce efficiently Biofilm formation
in the absence
of nanobacteria Sterile DMEM culture media will not form a biofilm on its own. When purified phosphotidyl inositol (a phospholipid common to biological membranes) was added to the culture, biofilm formation occurred within two weeks. The appearance of the particles was very similar to those found in nanobacterial cultures. The ability for a phospholipid to induce biofilm formation was prevented when the phospholipid was exposed to gamma radiation. The similar appearance between particles from those found in the sterile DMEM culture and the ones found in nanobacterial shows a high possibility that the nanobacteria could be made up of ingredients present in the DMEM media, and can take up a biological membrane within its structure. A combination between the ingredients of DMEM and the phospholipd allowed for growth. Since the biofilm that is created when the purified phosphotidyl inositol is added to the culture is similar to the particles from the nanobacteria, there is a high possibility that the nanobacteria have some traits that can define it as a living cell.
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